Evaluation of the influence of pre-analytical factors on total testosterone levels in healthy young men / Avaliação da influência de fatores pré-analíticos nas dosagens de testosterona total em homens jovens saudáveis

ABSTRACT Introduction: Testosterone is the main hormone that regulates male reproductive functions, directly participating in spermatogenesis and increasing sexual activity. The blood measurement of this hormone is essential for the diagnosis of neuroendocrine disorders, such as hypogonadism. However, there is lack of standardization regarding patient preparation for the hormone collection in clinical laboratories. Objective: Evaluate the effect of pre-analytical variables, including fasting, circadian and seasonal variation on testosterone levels in healthy young males. Material and methods: Forty-two volunteers were selected for the study, in the city of Barbacena, Minas Gerais. Four blood samples were collected from each of the participants, three in the winter: the first one in the morning after fasting; the second in the afternoon, without fasting; the third, in the next day morning, without fasting; and the last one taken in the summer, in the morning, after fasting. Results: The analyses showed that there was a significant decrease in total testosterone levels when there was no fasting for eight hours prior to collection and in the afternoon compared to the morning, both with p < 0.001. There was no significant difference in the results obtained in winter and summer. Conclusion: It is recommended that clinical laboratories standardize the collection of total testosterone by performing the test in the morning and after an eight-hour fast, in order to reduce variability and ensure reliability in the results.

RESUMEN Introducción: La testosterona es la principal hormona reguladora de funciones reproductoras masculinas, participando directamente en la espermatogénesis y en el aumento de la actividad sexual. La medición sanguínea de esa hormona es fundamental en el diagnóstico de trastornos neuroendocrinos, como el hipogonadismo. Sin embargo, hay poca estandarización en la preparación adecuada del paciente para la recolección de la hormona en laboratorios clínicos. Objetivo: Evaluar el efecto de variables preanalíticas, incluyendo ayuno, variación circadiana y estacional en las mediciones de testosterona en hombres jóvenes sanos. Material y métodos: Se eligieron 42 voluntarios para el estudio, en la ciudad de Barbacena, Minas Gerais. Se tomaron cuatro muestras de sangre de cada uno de los participantes, de las cuales tres en invierno: la primera, matutina, en ayunas; la segunda, vespertina, sin ayunas; la tercera, el dia siguiente, matutina, sin ayunas; el última se recolectó en verano, por la mañana, en ayunas. Resultados: Los análisis demostraron que hubo reducción significativa en los niveles de testosterona total cuando no se realizó el ayuno de ocho horas antes de la recolección y en el período vespertino en comparación al matutino, ambos con valor de p < 0,001. No hubo diferencia significativa en los resultados obtenidos en invierno y en verano. Conclusión: Se recomienda que los laboratórios clínicos estandarizen la recolección de testosterona total con la realización de la prueba en el período matutino y en ayuno de ocho horas, para reducir la variación y garantizar la confiabilidad de los resultados.

RESUMO Introdução: A testosterona é o principal hormônio regulador das funções reprodutivas masculinas, participando diretamente da espermatogênese e do aumento da atividade sexual. A dosagem sanguínea desse hormônio é fundamental no diagnóstico de distúrbios neuroendócrinos, como o hipogonadismo. Entretanto, há pouca padronização no preparo adequado do paciente para a coleta do hormônio em laboratórios clínicos. Objetivo: Avaliar o efeito de variáveis pré-analíticas, incluindo realização de jejum, variação circadiana e sazonal nas dosagens de testosterona em jovens saudáveis do sexo masculino. Material e métodos: Foram selecionados 42 voluntários para o estudo, na cidade de Barbacena, Minas Gerais. Quatro amostras de sangue de cada um dos participantes foram coletadas, sendo três no inverno: a primeira de manhã, em jejum; a segunda à tarde, sem jejum; a terceira no dia seguinte, de manhã, sem jejum. A última foi coletada no verão, na parte da manhã, em jejum. Resultados: As análises demonstraram que houve diminuição significativa dos níveis de testosterona total quando não foi realizado jejum de 8 horas antes da coleta e no período da tarde em comparação ao período da manhã, ambos com valor de p < 0,001. Não houve diferença significativa nos resultados obtidos no inverno e no verão. Conclusão: Recomendamos que os laboratórios clínicos padronizem a coleta de testosterona total com a realização do exame no período da manhã e em jejum de 8 horas, a fim de reduzir a variabilidade e garantir a confiabilidade nos resultados.

Synthesis of 2-ethylhexyl oleate catalyzed by Candida antarctica lipase immobilized on a magnetic polymer support in continuous flow.

This study investigated the synthesis of 2-ethylhexyl oleate catalyzed by Candida antarctica lipase immobilized on magnetic poly(styrene-co-divinylbenzene) particles in a continuous packed-bed bioreactor. Runs were carried out in a solvent-free system at 50 °C. The performance of the reactor was evaluated for substrates composed by oleic acid and 2-ethylhexanol at five molar ratios (1:4-4:1), determining its operation limits in terms of substrate flow rate. The system performance was quantified for three different flow rates corresponding to space-time between 3 and 12 h. For each condition, the influence of the space-time in the ester formation, esterification yield and productivity was determined. The molar ratio of acid-to-alcohol interfered, in a remarkable way, in the formation of 2-ethylhexyl oleate and the best performance was attained for substrate at equimolar ratio running at 12 h space-time. Under this condition, average 2-ethylhexyl oleate concentration was 471.65 ± 2.98 g L-1 which corresponded to ester productivity of 23.16 ± 0.49 mmol g-1 L-1 h-1. This strategy also gave high biocatalyst operational stability, revealing a half-life time of 2063 h. A model based on the ping-pong Bi-Bi mechanism was developed to describe the kinetics of the esterification reaction and validated using experimental data. The goodness of fit of the model was satisfactory (R2 = 0.9310-0.9952).

Continuous enzymatic biodiesel production from coconut oil in two-stage packed-bed reactor incorporating an extracting column to remove glycerol formed as by-product.

The transesterification of coconut oil with ethanol catalyzed by Burkholderia cepacia lipase immobilized on polysiloxane-polyvinyl alcohol was performed in a continuous flow. The experimental design consisted of a two-stage packed-bed reactor incorporating a column with cationic resin (Lewatit GF 202) to remove the glycerol formed as by-product and the reactor performance was quantified for three different flow rates corresponding to space-times from 10 to 14 h. The influence of space-time on the ethyl ester (FAEE) concentrations, yields and productivities was determined. The reactor operation was demonstrated for space-time of 14 h attaining FAEE concentrations of 58.5 ± 0.87 wt%, FAEE yields of 97.3 ± 1.9 % and productivities of 41.6  ± 1.0 mgester g medium (-1)  h(-1). Biodiesel purified samples showed average kinematic viscosity values of 5.5 ± 0.3 mm(2) s(-1) that meet the criteria established by the American National Standard ASTM (D6751). The immobilized lipase was found to be stable regarding its morphological and catalytic characteristics, showing half-life time (t 1/2) around 1540 h. The continuous packed-bed reactor connected in series with simultaneous glycerol removal has a great potential to attain high level of transesterification yields, raising biodiesel productivity.

Mucor circinelloides whole-cells as a biocatalyst for the production of ethyl esters based on babassu oil.

The intracellular lipase production by Mucor circinelloides URM 4182 was investigated through a step-by-step strategy to attain immobilized whole-cells with high lipase activity. Physicochemical parameters, such as carbon and nitrogen sources, inoculum size and aeration, were studied to determine the optimum conditions for both lipase production and immobilization in polyurethane support. Olive oil and soybean peptone were found to be the best carbon and nitrogen sources, respectively, to enhance the intracellular lipase activity. Low inoculum level and poor aeration rate also provided suitable conditions to attain high lipase activity (64.8 ± 0.8 U g(-1)). The transesterification activity of the immobilized whole- cells was assayed and optimal reaction conditions for the ethanolysis of babassu oil were determined by experimental design. Statistical analysis showed that M. circinelloides whole-cells were able to produce ethyl esters at all tested conditions, with the highest yield attained (98.1 %) at 35 °C using an 1:6 oil-to-ethanol molar ratio. The biocatalyst operational stability was also assayed in a continuous packed bed reactor (PBR) charged with glutaraldehyde (GA) and Aliquat-treated cells revealing half-life of 43.0 ± 0.5 and 20.0 ± 0.8 days, respectively. These results indicate the potential of immobilized M. circinelloides URM 4182 whole-cells as a low-cost alternative to conventional biocatalysts in the production of ethyl esters from babassu oil.

Effect of natural antioxidants on the lipase activity in the course of batch and continuous glycerolysis of babassu oil.

The effects of several natural antioxidants (copaiba oil, buriti oil, cocoa butter, tucuman butter, oregano and white thyme) were assessed in the enzymatic synthesis of monoglycerides (MAG) from the glycerolysis of babassu oil. The reactions were catalyzed by Burkholderia cepacia lipase immobilized on SiO2-PVA and the assays carried out in batch and continuous runs. Results were compared with those attained in the control reactions (without any strategy to avoid oxidation), and the best approach was tested in a continuous packed-bed reactor. The best performance was obtained using N2 in the reaction medium (60 % of MAG) followed by buriti oil (57.6 % of MAG) and cocoa butter (56.6 % of MAG), preventing the oxidation of babassu oil in batch reaction. However, the incorporation of buriti oil in the medium influenced the MAG profile, leading to the largest formation of monoolein, unlike other runs. Similar results were obtained in continuous reactions, using inert atmosphere and cocoa butter (24-25 % of MAG). Thereby, among the tested antioxidant agents, cocoa butter was the most effective in both systems, because it did not interfere in the MAG profile and also reduced the cost of the process.

Non-edible babassu oil as a new source for energy production-a feasibility transesterification survey assisted by ultrasound.

Ethyl esters of babassu oil were synthesized by alkaline catalysis to make the green production of biodiesel feasible with simple methods and available technology. Babassu oil is a transparent, light yellow oil extracted from the seeds of the babassu palm (Orbinya sp), and due to its high saturated fatty acid composition (83%), it is considered a non-inedible oil. Transesterification using ethanol represents a valid alternative to using methanol because of ethanol's lower toxicity and the higher yield on weight compared to methanol. Statistical methodology was applied to optimize the transesterification reaction, which was promoted by ultrasonic waves and mechanical agitation. Nuclear magnetic resonance spectroscopy was used to quantify the conversion attained. Alkaline transesterification assisted by ultrasound produced the best results with respect to reaction time and the phase separation step. The model obtained showed that conversions higher than 97% may be achieved in 10min with correct tuning of the process variables.

Evaluation of immobilized lipases on poly-hydroxybutyrate beads to catalyze biodiesel synthesis.

Five microbial lipase preparations from several sources were immobilized by hydrophobic adsorption on small or large poly-hydroxybutyrate (PHB) beads and the effect of the support particle size on the biocatalyst activity was assessed in the hydrolysis of olive oil, esterification of butyric acid with butanol and transesterification of babassu oil (Orbignya sp.) with ethanol. The catalytic activity of the immobilized lipases in both olive oil hydrolysis and biodiesel synthesis was influenced by the particle size of PHB and lipase source. In the esterification reaction such influence was not observed. Geobacillus thermocatenulatus lipase (BTL2) was considered to be inadequate to catalyze biodiesel synthesis, but displayed high esterification activity. Butyl butyrate synthesis catalyzed by BTL2 immobilized on small PHB beads gave the highest yield (≈90 mmol L(-1)). In biodiesel synthesis, the catalytic activity of the immobilized lipases was significantly increased in comparison to the free lipases. Full conversion of babassu oil into ethyl esters was achieved at 72 h in the presence of Pseudozyma antarctica type B (CALB), Thermomyces lanuginosus lipase (Lipex(®) 100 L) immobilized on either small or large PHB beads and Pseudomonas fluorescens (PFL) immobilized on large PHB beads. The latter preparation presented the highest productivity (40.9 mg of ethyl esters mg(-1) immobilized protein h(-1)).

Immobilization of a Commercial Lipase from Penicillium camembertii (Lipase G) by Different Strategies.

The objective of this work was to select the most suitable procedure to immobilize lipase from Penicillium camembertii (Lipase G). Different techniques and supports were evaluated, including physical adsorption on hydrophobic supports octyl-agarose, poly(hydroxybutyrate) and Amberlite resin XAD-4; ionic adsorption on the anionic exchange resin MANAE-agarose and covalent attachment on glyoxyl-agarose, MANAE-agarose cross-linked with glutaraldehyde, MANAE-agarose-glutaraldehyde, and epoxy-silica-polyvinyl alcohol composite. Among the tested protocols, the highest hydrolytic activity (128.2 ± 8.10 IU·g(-1) of support) was achieved when the lipase was immobilized on epoxy-SiO(2)-PVA using hexane as coupling medium. Lipase immobilized by ionic adsorption on MANAE-agarose also gave satisfactory result, attaining 55.6 ± 2.60 IU·g(-1) of support. In this procedure, the maximum loading of immobilized enzyme was 9.3 mg·g(-1) of gel, and the highest activity (68.8 ± 2.70 IU·g(-1) of support) was obtained when 20 mg of protein·g(-1) was offered. Immobilization carried out in aqueous medium by physical adsorption on hydrophobic supports and covalent attachment on MANAE-agarose-glutaraldehyde and glyoxyl-agarose was shown to be unfeasible for Lipase G. Thermal stability tests revealed that the immobilized derivative on epoxy-SiO(2)-PVA composite using hexane as coupling medium had a slight higher thermal stability than the free lipase.

Análise microscópica do miocárdio ventricular esquerdo em cães soropositivos para cinomose / Microscopic analysis of the left ventricular myocardium in positive serum dogs to distemper disease

Classificado no gênero Morbillivirus da família Paramixoviridae, o vírus da cinomose possui RNA de fita simples de polaridade negativa, é causador de doença multissistêmica, altamente contagiosa e grave dos cães e carnívoros selvagens, e com elevado índice de mortalidade em animais não vacinados ou com falhas vacinais. Com o objetivo de avaliar as alterações histopatológicas no coração, particularmente na região do miocárdio ventricular esquerdo, de cães naturalmente infectados com o vírus da cinomose, foram estudados 35 animais, de ambos os sexos e com idades variadas. Das 35 amostras enviadas ao Laboratório de Medicina Veterinária Preventiva do Hospital Veterinário de Uberaba, 100 por cento (35/35) mostrou-se soropositivas para a cinomose (técnica de imunoensaio em fase sólida) e tiveram no miocárdio ventricular esquerdo as seguintes alterações histopatológicas: miocardite, degeneração hialina, hiperemia e hemorragia, com 42,8 por cento (15/35), 31,4 por cento (11/35), 14,3 por cento (5/35) e 11,4 por cento (4/35), respectivamente. Tendo utilizado o teste Qui-Quadrado com nível de significância de 0,05, conclui-se que existe alta correlação (p=0,02) entre os animais infectados com o vírus da cinomose e as alterações histopatológicas observadas no miocárdio ventricular esquerdo.

Classified pertaining to the genus Morbillivirus of the Paramyxoviridae family, the canine distemper virus is a RNA single-stranded virus with negative polarity and causes a multisystemic disease, serious and highly contagious for dogs and wild carnivores, with a high mortality rate in non-vaccinated animals or with vaccine fails. With the objective to evaluate heart histopathological alterations, particularly in the left ventricular myocardium, in dogs naturally infected with canine distemper virus, 35 dogs, males and females of different ages, were studied. All the 35 samples sent to the Veterinary Hospital of Uberaba were serum-positive for distemper (immunoassay technique in solid phase) and had in the left ventricular myocardium the following histopathologic alterations: myocarditis, hyalin degeneration, hyperemia and hemorrhage, in 42.8 percent (15/35), 31.4 percent (11/35), 14.3 percent (5/35) and 11.4 percent (4/35), respectively. Having carried out the Qui-Quadrado test with a significancy level of 0.05, it can be concluded that there is a high correlation (p=0.02) between the infected animals with canine distemper virus and histopathological alterations found in the left ventricular myocardium.